This application is based on applications NO.HEI10-249731 filed in Japan, the content of which is incorporated hereinto by reference.
1. Field of the Invention
This invention relates to a reference electrode used for electrochemically determining a particular chemical substance in a solution via a reaction such as an enzyme reaction as well as a biosensor therewith.
2. Description of the Related Art
The structure of a conventional reference electrode will be described with reference to FIG. 4, consisting of a cross-section and a plan view for a reference electrode disclosed in JP-A 9-182738. In FIG. 4(a), a polyimide layer PI1, an anode film M and a silver layer S are sequentially formed on a cathode film K. The silver layer S acts as a reference electrode, on which the polyimide layer PI2 is formed as a protection layer. It is described that instead of the polyimide layer PI2, a water-repelling photoresist may be formed as a protection layer. It is also described that such a protection layer can be formed on a reference electrode to extend the life of the electrode.
It has been, however, difficult to achieve a sufficiently long life for the above reference electrode because a part of the surface of the reference electrode is directly in contact with an electrolyte, so that silver chloride formed on the surface is likely to be dissociated in the electrolyte. According to our examination, a reference electrode having the above structure can usually operate for about 2 days while maintaining its initial performance.
When a water-repelling photoresist is used as a material for the protection layer in the above reference electrode, it may make a manufacturing process more complicated, leading to increase in a production cost. Thus, the technique has a room for improvement. It is because formation of a photoresist requires repeating exposure and development using a photolithography technique and forming an electrolyte injection port for injecting an electrolyte and open holes for removing bubbles generated during injection of the electrolyte.
Furthermore, there has been a problem that when the above reference electrode is applied to a biosensor, contaminants in a specimen adhere to its surface, causing reduction in sensitivity. The problem will be described.
A biosensor utilizes a catalytic enzyme to convert chemical substances in a solution into, e.g., hydrogen peroxide, which is then determined using an oxidation-reduction reaction. It is important to eliminate effects of interferent materials and contaminants for achieving stable properties and an extended life in a biosensor. An interferent material refers to a chemical substance which may affect the above oxidation-reduction reaction system to give a positive error in a measurement result, such as ascorbic acid and acetaminophen. A contaminant refers to a chemical substance which may be adsorbed by an electrode surface to give a negative error in a measurement result, such as albumin, urea, urea compounds and creatinine.
The above reference electrode according to the prior art, however, uses polyimide and/or a photoresist as a material for a protection layer and thus contaminants may easily attach to the surface of the protection layer. It has been, therefore, difficult to avoid reduction in sensitivity after a long-term use.
This invention provides a reference electrode comprising an electrode on an insulating substrate and a protection layer covering the electrode, said protection layer mainly consisting of a polymer in which a pendant group containing at least a fluoroalkylene block is attached to a non-fluorinated vinyl polymer.
That is, a reference electrode according to this invention consists of an electrode (electrode layer) on an insulating substrate and multiple layers having different functions on the electrode.
A reference electrode herein refers to an electrode as a standard for determining an electrode potential of a working electrode. The reference electrode of this invention may be suitably used for electrochemically determining a particular chemical substance in a solution using an enzyme electrode and thus suitably used for a part of, e.g., a biosensor.
In this invention, the polymer composing the protection layer has a pendant group comprising a fluoroalkylene block (fluoroalkylene unit). Such a configuration may prevent adhesion of contaminants such as proteins and urea compounds to provide a reference electrode exhibiting stable output properties even for a long-term use. The fluoroalkylene moiety may not contribute to dissolution into a washing agent such as non-fluorinated solvent and a surfactant, providing a reference electrode with good chemical resistance.
The polymer has a non-fluorinated vinyl polymer structure as a principal chain, which is highly adhesive to an electrode or another organic polymer layer formed on the electrode. It, therefore, does not cause a gap between the electrode or a layer such as an organic polymer layer formed on the electrode surface and the protection layer. Therefore, the electrode surface is not in contact with an electrode, which prevents metal composing the electrode portion from eluting into the electrode and leads to reduction in a time for washing the reference electrode.
In addition, its good adhesiveness to, e.g., the electrode may improve durability of the layered structure to provide a reference electrode resistant to deterioration due to a long-term use.
The polymer may have, in addition to the pendant group comprising a fluoroalkylene block, any other appropriate side chain or functional group; for example, a properly polar functional group such as xe2x80x94OH and xe2x80x94COOH groups may further improve adhesiveness to an electrode or another organic polymer layer formed on the electrode.
The protection layer in this invention may be formed as a homogeneous film by a convenient process such as dip-coating, spin-coating and spray-coating and may be suitable for mass production.
This invention also provides a reference electrode comprising an electrode on an insulating substrate and a protection layer covering the electrode, said protection layer mainly consisting of a polycarboxylic acid (A) fluoroalcohol ester.
This invention also provides a reference electrode comprising an electrode on an insulating substrate and a protection layer covering the electrode, said protection layer mainly consisting of a polycarboxylic acid (A) fluoroalcohol ester and a polycarboxylic acid (B) alkylalcohol ester.
This invention also provides a reference electrode comprising an electrode on an insulating substrate and a protection layer covering the electrode, said protection layer mainly consisting of a polycarboxylate comprising alkylalcohol ester and fluoroalcohol ester groups.
These reference electrodes consist of an electrode (electrode layer) on an insulating substrate and multiple layers having different functions on the electrode, characterized in that the protection layer is composed of a polymer having a particular structure.
In a reference electrode according to this invention, a fluoroalcohol polycarboxylate is used as a material for a protection layer. A fluoroalcohol polycarboxylate refers to a polycarboxylic acid, whose carboxyl groups are partially or totally esterified with a fluoroalcohol. A fluoroalcohol refers to an alcohol, at least one or all of whose hydrogen atoms are replaced with fluorine atom(s).
The protection layer material has a fluoroalcohol ester group, which may prevent adhesion of contaminants such as proteins and urea compounds, leading to a reference electrode exhibiting stable properties when used for a long term. The fluoroalcohol ester group makes the material insoluble in almost any non-fluorinated solvent or a washing agent such as a surfactant, to give a reference electrode with an improved chemical resistance.
The reference electrode of this invention has a protection layer comprising a polymer having a principal chain of a polycarboxylic acid, to which a fluoroalcohol is attached via an ester group. It, therefore, can improve adhesiveness to an electrode or another organic polymer layer formed on the electrode and prevent causing a gap between the electrode or the organic polymer layer and the protection layer. Therefore, the electrode surface is not in contact with an electrode, which prevents metal composing the electrode portion from eluting into the electrode and leads to reduction in a time for washing the reference electrode.
In addition, its good adhesiveness to, e.g., the electrode may improve durability of the layered structure to provide a reference electrode resistant to deterioration due to a long-term use.
The polymer may have, in addition to the fluoroalcohol ester group, any other appropriate functional group to the principal chain; a properly polar functional group may further improve adhesiveness to another organic polymer layer such as an adjacent electrode or a layer such as an organic polymer layer formed on the electrode.
When a protection layer comprises (a) a polycarboxylic acid (A) fluoroalcohol ester and a polycarboxylic acid (B) alkylalcohol ester or (b) a polycarboxylate having an alkylalcohol ester and a fluoroalcohol ester groups, high temperature stability may be improved, in addition to the above effects. A reference electrode or a biosensor therewith may be sometimes stored or used at a relatively higher temperature (for example, ca. 40xc2x0 C.). When used for measurement after leaving at a higher temperature, sensitivity of a conventional reference electrode has often varied significantly, compared to measurement before exposure to the higher temperature. On the other hand, a reference electrode or biosensor comprising the above protection layer little varies in its sensitivity even after exposure to a higher temperature.
In a reference electrode of this invention as described above, an electrode and a protection layer may be formed in direct contact with each other or another layer may intervene these layers. For example, a binding layer mainly consisting of a silane-coupling agent is disposed between the electrode and the protection layer.
The protection layer may be formed as a homogeneous film by a convenient process such as dipping, spin-coating and spray coating and may be suitable for mass production.
This invention also provides a biosensor using the above reference electrode. The biosensor has a protection layer comprising the polymer having the above particular structure on the reference electrode surface. It, therefore, may be excellent in long-term stability and may be used under a wide range of measuring conditions.
This invention also relates to a variety of measuring apparatus using the above biosensor. Specifically, this invention provides a measuring apparatus comprising the above biosensor and a data indicator indicating an electric signal from the biosensor.
This invention also provides a measuring apparatus comprising the above biosensor, an electrochemical measuring circuit receiving an electric signal from the biosensor, a data processor calculating a measured value based on the electric signal and a data indicator indicating the measured value.
These measuring apparatuses may realize excellent long-term stability and may be used under wide ranges of measuring conditions because of their biosensor having a particular structure of working electrode. They are also easily operated even by an unfamiliar individual.
This invention also provides a method for manufacturing a reference electrode comprising the steps of: forming an electrode on an insulating substrate; and applying a liquid containing a polymer in which a pendant group having at least a fluoroalkylene block is attached to a non-fluorinated vinyl polymer, to the electrode directly or via another layer and then drying it to form a protection layer.
In this manufacturing process, a protection layer is formed by applying and then drying a liquid comprising a polymer having the above particular structure. Thus, there may be provided, with a good controllability for a film thickness, a protection layer which is excellent in stability for repeated measurement, adhesiveness to adjacent layers and durability. Since the liquid containing the above polymer has a lower viscosity, the protection layer may be readily formed with a reduced film thickness. Specifically, a protection layer 0.01 to 3 xcexcm of thickness after drying may be satisfactorily formed.